1. Field of the Invention
This invention relates to a composition for measuring intravital ingredients such as glucose, ATP, creatine kinase, other phosphotransferases, various glucose-releasing glycosidases, and the like.
2. Description of the Prior Art
In clinical testing for the measurement of glucose or creatine kinase (hereinafter abbreviated CPK), the usual analytical method employed comprises a coupling enzyme system of hexokinase (hereinafter abbreviated as HK) and glucose-6-phosphate dehydrogenase (hereinafter abbreviated G6PDH). This system is referred to as the HK/G6PDH system, and has been extensively used because of the high availability of HK.
The principle of the system can be illustrated by the following reaction scheme: ##STR1##
In the above reaction sequence, CP represents creatine phosphate, C represents creatine, ADP represents adenosine 5'-diphosphate, ATP represents adenosine 5'-triphosphate, G6P represents glucose-6-phosphate, NAD(P) represents nicotinamidoadeninedinucleotide (phosphate), NAD(P)H represents reduced nicotinamidoadeninedinucleotide (phosphate), and 6-PGA represents 6-phosphogluconic acid. The enzymes which catalyze reactions (1), (2) and (3) are CPK, HK, and G6PDH, respectively.
HK, however, has the drawback that it literally acts on many hexose and phosphorylate sugars other than glucose, such as fructose and mannose, which exist in the intravital liquids. HK of yeast origin which is commonly used in the KH/G6PDH system shows a 1.8 times higher reaction rate for fructose than for glucose. Each of these phosphorylated hexoses is finally converted to G6P by the action of phosphomannose isomerase (hereinafter abbreviated as PMI) or phosphoglucose isomerase (hereinafter abbreviated as PGI) which are also present within intravital liquids by the scheme shown below which results in a positive error of the measured value. ##STR2## In the above scheme, M6P represents mannose-6-phosphate, and F6P represents fructose-6-phosphate.
As a result of an intensive study whose objective has been to eliminate the drawbacks of the HK/G6PDH system, it has been discovered that glucokinase (hereinafter abbreviated as GK) has a high substrate specificity for glucose as a phosphotransferase and thus can act as a substitute for HK. This discovery is the basis of the present invention.